Genetic changes causing brain size expansion in human evolution have remained elusive. Notch signaling is essential for radial glia stem cell proliferation and is a determinant of neuronal number in ...the mammalian cortex. We find that three paralogs of human-specific NOTCH2NL are highly expressed in radial glia. Functional analysis reveals that different alleles of NOTCH2NL have varying potencies to enhance Notch signaling by interacting directly with NOTCH receptors. Consistent with a role in Notch signaling, NOTCH2NL ectopic expression delays differentiation of neuronal progenitors, while deletion accelerates differentiation into cortical neurons. Furthermore, NOTCH2NL genes provide the breakpoints in 1q21.1 distal deletion/duplication syndrome, where duplications are associated with macrocephaly and autism and deletions with microcephaly and schizophrenia. Thus, the emergence of human-specific NOTCH2NL genes may have contributed to the rapid evolution of the larger human neocortex, accompanied by loss of genomic stability at the 1q21.1 locus and resulting recurrent neurodevelopmental disorders.
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•NOTCH2NLA,B,C expressed in human fetal brain radial glia stem cells, arose 0.5–4 mya•These genes encode secreted, Notch-related proteins that enhance Notch signaling•Overexpression of NOTCH2NL delays neuronal differentiation, while deletion accelerates it•NOTCH2NLA and NOTCH2NLB serve as breakpoints in 1q21.1 deletion/duplication syndrome
Human-specific Notch paralogs are expressed in radial glia, enhance Notch signaling, and impact neuronal differentiation.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP
The advent of induced pluripotent stem cell (iPSC)-derived neurons has revolutionized Parkinson’s disease (PD) research, but single-cell transcriptomic analysis suggests unresolved cellular ...heterogeneity within these models. Here, we perform the largest single-cell transcriptomic study of human iPSC-derived dopaminergic neurons to elucidate gene expression dynamics in response to cytotoxic and genetic stressors. We identify multiple neuronal subtypes with transcriptionally distinct profiles and differential sensitivity to stress, highlighting cellular heterogeneity in dopamine in vitro models. We validate this disease model by showing robust expression of PD GWAS genes and overlap with postmortem adult substantia nigra neurons. Importantly, stress signatures are ameliorated using felodipine, an FDA-approved drug. Using isogenic SNCA-A53T mutants, we find perturbations in glycolysis, cholesterol metabolism, synaptic signaling, and ubiquitin-proteasomal degradation. Overall, our study reveals cell type-specific perturbations in human dopamine neurons, which will further our understanding of PD and have implications for cell replacement therapies.
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•Deconvolution of in vitro human dopamine neuron heterogeneity by scRNA-seq•Identification of dopamine subtypes with transcriptionally distinct profiles•Cell type-specific gene perturbations of cholesterol metabolism in stress response•PD GWAS associated gene expression analysis at the single-cell level
Fernandes et al. identify and characterize cellular heterogeneity in human iPSC dopamine neurons using single-cell RNA-seq. The different subtypes of dopamine neurons illustrate distinct transcriptional profiles in response to toxic and genetic stressors. Furthermore, PD GWAS associated gene expression analysis is performed at the single-cell level.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP
IMPORTANCE: Although asthma is a chronic disease, the expected rate of spontaneous remissions of adult asthma and the stability of diagnosis are unknown. OBJECTIVE: To determine whether a diagnosis ...of current asthma could be ruled out and asthma medications safely stopped in randomly selected adults with physician-diagnosed asthma. DESIGN, SETTING, AND PARTICIPANTS: A prospective, multicenter cohort study was conducted in 10 Canadian cities from January 2012 to February 2016. Random digit dialing was used to recruit adult participants who reported a history of physician-diagnosed asthma established within the past 5 years. Participants using long-term oral steroids and participants unable to be tested using spirometry were excluded. Information from the diagnosing physician was obtained to determine how the diagnosis of asthma was originally made in the community. Of 1026 potential participants who fulfilled eligibility criteria during telephone screening, 701 (68.3%) agreed to enter into the study. All participants were assessed with home peak flow and symptom monitoring, spirometry, and serial bronchial challenge tests, and those participants using daily asthma medications had their medications gradually tapered off over 4 study visits. Participants in whom a diagnosis of current asthma was ultimately ruled out were followed up clinically with repeated bronchial challenge tests over 1 year. EXPOSURE: Physician-diagnosed asthma established within the past 5 years. MAIN OUTCOMES AND MEASURES: The primary outcome was the proportion of participants in whom a diagnosis of current asthma was ruled out, defined as participants who exhibited no evidence of acute worsening of asthma symptoms, reversible airflow obstruction, or bronchial hyperresponsiveness after having all asthma medications tapered off and after a study pulmonologist established an alternative diagnosis. Secondary outcomes included the proportion with asthma ruled out after 12 months and the proportion who underwent an appropriate initial diagnostic workup for asthma in the community. RESULTS: Of 701 participants (mean SD age, 51 16 years; 467 women 67%), 613 completed the study and could be conclusively evaluated for a diagnosis of current asthma. Current asthma was ruled out in 203 of 613 study participants (33.1%; 95% CI, 29.4%-36.8%). Twelve participants (2.0%) were found to have serious cardiorespiratory conditions that had been previously misdiagnosed as asthma in the community. After an additional 12 months of follow-up, 181 participants (29.5%; 95% CI, 25.9%-33.1%) continued to exhibit no clinical or laboratory evidence of asthma. Participants in whom current asthma was ruled out, compared with those in whom it was confirmed, were less likely to have undergone testing for airflow limitation in the community at the time of initial diagnosis (43.8% vs 55.6%, respectively; absolute difference, 11.8%; 95% CI, 2.1%-21.5%). CONCLUSIONS AND RELEVANCE: Among adults with physician-diagnosed asthma, a current diagnosis of asthma could not be established in 33.1% who were not using daily asthma medications or had medications weaned. In patients such as these, reassessing the asthma diagnosis may be warranted.
RNA polymerase II (RNAPII) pausing in early elongation is critical for gene regulation. Paused RNAPII can be released into productive elongation by the kinase P-TEFb or targeted for premature ...termination by the Integrator complex. Integrator comprises endonuclease and phosphatase activities, driving termination by cleavage of nascent RNA and removal of stimulatory phosphorylation. We generated a degron system for rapid Integrator endonuclease (INTS11) depletion to probe the direct consequences of Integrator-mediated RNA cleavage. Degradation of INTS11 elicits nearly universal increases in active early elongation complexes. However, these RNAPII complexes fail to achieve optimal elongation rates and exhibit persistent Integrator phosphatase activity. Thus, only short transcripts are significantly upregulated following INTS11 loss, including transcription factors, signaling regulators, and non-coding RNAs. We propose a uniform molecular function for INTS11 across all RNAPII-transcribed loci, with differential effects on particular genes, pathways, or RNA biotypes reflective of transcript lengths rather than specificity of Integrator activity.
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•Rapid degradation of INTS11 globally increases early elongation complexes•In the absence of INTS11, RNAPII released into genes is deficient in elongation•Loss of INTS11 endonuclease does not disrupt Integrator-PP2A phosphatase function•Predominantly short genes and non-coding RNAs are upregulated upon INTS11 degradation
Stein et al. show that Integrator broadly balances levels of RNAPII pause release with premature termination. Acute depletion of Integrator endonuclease allows for increased RNAPII release into coding and non-coding RNAs. However, RNAPII elongation is inefficient, and only short RNAs are upregulated, including several transcription factors that initiate signaling pathways.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP
Precise regulation of transcription by RNA polymerase II (RNAPII) is critical for organismal growth and development. However, what determines whether an engaged RNAPII will synthesize a full-length ...transcript or terminate prematurely is poorly understood. Notably, RNAPII is far more susceptible to termination when transcribing non-coding RNAs than when synthesizing protein-coding mRNAs, but the mechanisms underlying this are unclear. To investigate the impact of transcribed sequence on elongation potential, we developed a method to screen the effects of thousands of INtegrated Sequences on Expression of RNA and Translation using high-throughput sequencing (INSERT-seq). We found that higher AT content in non-coding RNAs, rather than specific sequence motifs, drives RNAPII termination. Further, we demonstrate that 5' splice sites autonomously stimulate processive transcription, even in the absence of polyadenylation signals. Our results reveal a potent role for the transcribed sequence in dictating gene output and demonstrate the power of INSERT-seq toward illuminating these contributions.
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EMUNI, FIS, FZAB, GEOZS, GIS, IJS, IMTLJ, KILJ, KISLJ, MFDPS, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, SBMB, SBNM, UKNU, UL, UM, UPUK, VKSCE, ZAGLJ
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Overdiagnosis of asthma in obese and nonobese adults Aaron, Shawn D., MD; Vandemheen, Katherine L., BScN; Boulet, Louis-Philippe, MD ...
CMAJ. Canadian Medical Association journal,
11/2008, Volume:
179, Issue:
11
Journal Article
Peer reviewed
Open access
It is unclear whether asthma is overdiagnosed in developed countries, particularly among obese individuals, who may be more likely than nonobese people to experience dyspnea.
We conducted a ...longitudinal study involving nonobese (body mass index 20-25) and obese (body mass index >/= 30) individuals with asthma that had been diagnosed by a physician. Participants were recruited from 8 Canadian cities by means of random-digit dialing. A diagnosis of current asthma was excluded in those who did not have evidence of acute worsening of asthma symptoms, reversible airflow obstruction or bronchial hyperresponsiveness, despite being weaned off asthma medications. We stopped asthma medications in those in whom a diagnosis of asthma was excluded and assessed their clinical outcomes over 6 months.
Of 540 individuals with physician-diagnosed asthma who participated in the study, 496 (242 obese and 254 nonobese) could be conclusively assessed for a diagnosis of asthma. Asthma was ultimately excluded in 31.8% (95% confidence interval CI 26.3%-37.9%) in the obese group and in 28.7% (95% CI 23.5%-34.6%) in the nonobese group. Overdiagnosis of asthma was no more likely to occur among obese individuals than among nonobese individuals (p = 0.46). Of those in whom asthma was excluded, 65.5% did not need to take asthma medication or seek health care services because of asthma symptoms during a 6-month follow-up period.
About one-third of obese and nonobese individuals with physician-diagnosed asthma did not have asthma when objectively assessed. This finding suggests that, in developed countries such as Canada, asthma is overdiagnosed.
Approximately 5-10% of adults may have undiagnosed airflow obstruction. The objective of this study was to develop a population-based case-finding strategy to assess the prevalence of undiagnosed ...airflow obstruction (asthma or COPD) amongst adults with respiratory symptoms in Canada.
Adults without a previous history of asthma, COPD, or lung disease were recruited by random digit-dialling and asked if they had symptoms of dyspnea, cough, sputum or wheeze within the past 6 months. Those who answered affirmatively completed the Asthma Screening Questionnaire (ASQ), COPD-Diagnostic Questionnaire (COPD-DQ), and COPD Assessment Test (CAT). Those with ASQ score ≥6 or COPD-DQ score ≥20 underwent pre and post bronchodilator spirometry to diagnose asthma or COPD.
12 117 individuals were contacted at home and assessed for study eligibility. Of 1260 eligible individuals, 910 (72%) enrolled and underwent spirometry. Ultimately, 184 subjects (20% of those enrolled) had obstructive lung disease (73 asthma and 111 COPD). Individuals found to have undiagnosed asthma or COPD had more severe respiratory symptoms and impaired quality of life compared to those without airflow obstruction. The ASQ, COPD-DQ, and CAT had ROC areas for predicting undiagnosed asthma or COPD of 0.49, 0.64 and 0.56 respectively. Four descriptive variables; age, BMI, sex and pack-years smoked, produced better ROC values than the questionnaires (ROC area=0.68).
Twenty percent of randomly-selected individuals who report respiratory symptoms in Canada have undiagnosed airflow obstruction due to asthma or COPD. Questionnaires could exclude subjects at low risk but lacked ability to accurately find subjects with undiagnosed disease.
In patients with a history suggestive of asthma, diagnosis is usually confirmed by spirometry with bronchodilator response (BDR) or confirmatory methacholine challenge testing (MCT).
We examined the ...proportion of participants with negative BDR testing who had a positive MCT (and its predictors) result and characteristics of MCT, including effects of controller medication tapering and temporal variability (and predictors of MCT result change), and concordance between MCT and pulmonologist asthma diagnosis.
Adults with self-reported physician-diagnosed asthma were recruited by random-digit dialing across Canada. Subjects performed spirometry with BDR testing and returned for MCT if testing was nondiagnostic for asthma. Subjects on controllers underwent medication tapering with serial MCTs over 3 to 6 weeks. Subjects with a negative MCT (the provocative concentration of methacholine that results in a 20% drop in FEV1 PC20 > 8 mg/mL) off medications were examined by a pulmonologist and had serial MCTs after 6 and 12 months.
Of 500 subjects (50.5 ± 16.6 years old, 68.0% female) with a negative BDR test for asthma, 215 (43.0%) had a positive MCT. Subjects with prebronchodilator airflow limitation were more likely to have a positive MCT (OR, 1.90; 95% CI, 1.17-3.04). MCT converted from negative to positive, with medication tapering in 18 of 94 (19.1%) participants, and spontaneously over time in 25 of 165 (15.2%) participants. Of 231 subjects with negative MCT, 28 (12.1%) subsequently received an asthma diagnosis from a pulmonologist.
In subjects with a self-reported physician diagnosis of asthma, absence of bronchodilator reversibility had a negative predictive value of only 57% to exclude asthma. A finding of spirometric airflow limitation significantly increased chances of asthma. MCT results varied with medication taper and over time, and pulmonologists were sometimes prepared to give a clinical diagnosis of asthma despite negative MCT. Correspondingly, in patients for whom a high clinical suspicion of asthma exists, repeat testing appears to be warranted.
The cerebral cortex has expanded in size and complexity in primates, yet the molecular innovations that enabled primate-specific brain attributes remain obscure. We generated cerebral cortex ...organoids from human, chimpanzee, orangutan, and rhesus pluripotent stem cells and sequenced their transcriptomes at weekly time points for comparative analysis. We used transcript structure and expression conservation to discover gene regulatory long non-coding RNAs (lncRNAs). Of 2,975 human, multi-exonic lncRNAs, 2,472 were structurally conserved in at least one other species and 920 were conserved in all. Three hundred eighty-six human lncRNAs were transiently expressed (TrEx) and many were also TrEx in great apes (46%) and rhesus (31%). Many TrEx lncRNAs are expressed in specific cell types by single-cell RNA sequencing. Four TrEx lncRNAs selected based on cell-type specificity, gene structure, and expression pattern conservation were ectopically expressed in HEK293 cells by CRISPRa. All induced trans gene expression changes were consistent with neural gene regulatory activity.
•New orangutan and chimpanzee iPSC lines enable pan-primate gene expression analysis•Transiently expressed (TrEx) lncRNAs identified in primate cerebral cortex organoids•Conserved TrEx pattern correlates with cell-type specificity by single-cell RNA-seq•Four primate-conserved TrEx lncRNAs influence neural genes when ectopically expressed
In this article, Salama and colleagues identified transiently expressed (TrEx) lncRNAs from human, chimpanzee, orangutan, and rhesus pluripotent stem cell-derived cerebral cortex organoids and assessed their structural and expression conservation. Transient expression correlated with cell-type specificity as measured by single-cell RNA-seq. Ectopic expression of TrEx lncRNAs by CRISPRa modulated expression of neural genes in trans, suggesting regulatory function.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP